Epizootiologa, prevencin y control de la coriza infecciosa

Epizootiology, prevention and control of infectious coryza

Edgardo Soriano Vargas*
Horacio Ral Terzolo**

Abstract
In the present paper, the epizootiology of infectious coryza, an upper respiratory tract disease of poultry
is reviewed. The disease is characterized by sneezing, nasal discharge and facial swelling. However, very
virulent strains have also been described as causing lesions of pneumonia, airsacculitis and arthritis.
Pathogenic mechanisms and virulence factors of etiologic agent, Haemophilus paragallinarum, identified
at the date, are described. Furthermore, particular emphasis is made concerning the diagnosis of the
disease and identification of the causal agent. Also, prevention and control strategies of the disease are
reviewed.
Key words: INFECTIOUS CORYZA, H AEMOPHILUS PARAGALLINARUM , POULTRY DISEASES, CHICKENS.

Resumen
Aqu se revisa la epizootiologa de la coriza infecciosa, enfermedad del tracto respiratorio superior de
los pollos, que se caracteriza por estornudo, descarga nasal e inflamacin facial. Se han descrito cepas
muy virulentas que causan lesiones de neumona, aerosaculitis y artritis. Se describen los mecanismos
de patogenicidad y virulencia de Haemophilus paragallinarum. Adems, se enfatiza el diagnstico de la
enfermedad e identificacin del agente causal, y se revisan estrategias de prevencin y control de la
enfermedad.
Palabras clave: CORIZA INFECCIOSA, H AEMOPHILUS
POLLOS.

PARAGALLINARUM,

ENFERMEDADES DE LAS AVES,

Recibido para su publicacin el 11 de diciembre de 2003 y aceptado el 3 de junio del 2004.

*Programa de Doctorado en Ciencias de la Produccin y de la Salud Animal, Facultad de Medicina Veterinaria y Zootecnia,
Universidad Nacional Autnoma de Mxico, 04510, Mxico, D. F. Direccin actual: Centro de Investigacin y Estudios
Avanzados en Salud Animal, Facultad de Medicina Veterinaria y Zootecnia, Universidad Autnoma del Estado de Mxico,
50000, Toluca, Estado de Mxico, Mxico. E-mail: soriano@uaemex.mx
**
Instituto Nacional de Tecnologa Agropecuaria, CC 276, 7620, Balcarce, Argentina. E-mail: terzolo@balcarce.inta.gov.ar

Vet. Mx., 35 (3) 2004

261

Introduction

Introduccin

a coriza infecciosa es una enfermedad del tracto

respiratorio superior de los pollos, se caracteriza por producir descarga nasal, estornudo
e inflamacin facial. El agente etiolgico de esta
enfermedad es la bacteria Haemophilus paragallinarum . El
impacto econmico de esta enfermedad radica en las
prdidas que ocasiona a la avicultura, debido a retraso
del crecimiento, prdida de peso, incremento en el
nmero de aves eliminadas y predisposicin a la enfermedad respiratoria crnica complicada. En gallinas
de postura, la produccin de huevo puede reducirse
considerablemente.1

nfectious coryza is an upper respiratory tract disease of chickens, characterized by producing nasal
discharge, sneezing, and facial swelling. The bacterium Haemophilus paragallinarum is the etiologic agent
of the disease. Economic impact of disease result from
losses in poultry, due to growth retardation, increased
number of culls and predisposing to chronic complicated disease. Egg production is considerably drops in
laying flocks.1

Epizootiology
Natural and experimental hosts
Chicken (Gallus gallus) is the natural host for H.
paragallinarum and birds of all ages are susceptible.1
It has been reported that this bacterium was isolated
from quail 2,3 and some psittacines. 4,5 In addition
signs of coryza and sinusitis were observed in three
experimentally infected turkeys, these lesions were
similar to the ones observed in chickens inoculated
with the same bacterial culture. 6 However, other
bacteriological conclusive studies showing evidence the
presence and susceptibility to Haemophilus paragallinarum
in other avian species have not been carried out.
Rabbits, guinea pigs, mice, sparrows, ducks7 and
pigeons6 are refractory to experimental infection.

Transmission, carriers, and vectors

The main reservoir of infection is the chicken itself,
chronic or healthy carrier birds are both involved. 8
Infectious coryza outbreaks are more frequent during
the fall and winter. 9 Sparrows (Paser paser) have
implicating as vectors. Some epidemiological studies
suggest air-borne transmission as a possible mechanism
for introducing this microorganism into isolated
farms.10

Incubation period
The incubation period of infectious coryza varies
between 24 to 48 h after experimental inoculation
of susceptible birds with an live culture of H.
paragallinarum or infectious exudates. The incubation
period may be variable according to each particular
experimental exposure condition: 24 h, intrasinusal
inoculation; 48 h, nasal instillation; 72 h, birds in
cages; 4 days, infected-water contact, and 6 to 14 days
by air-borne transmission.11

262

Epizootiologa
Hospederos naturales y experimentales
Los pollos y gallinas (Gallus gallus) son hospederos
naturales de H. paragallinarum, susceptibles en todas
las edades.1 No obstante, existen informes del
aislamiento de esta bacteria en codornices 2,3 y
psitcidos. 4,5 Tres pavos mostraron signos de coriza y
sinusitis similares a los observados en pollos desafiados
experimentalmente con el mismo cultivo. 6 Sin
embargo, no se han efectuado estudios bacteriolgicos
definitivos que evidencien presencia y susceptibilidad
de otras especies aviares a H. paragallinarum. Los
conejos, cobayos, ratones, gorriones, patos7 y palomas 6
son refractarios a la infeccin experimental.

Transmisin, portadores y vectores

Los principales reservorios de infeccin son aves con
infeccin crnica y portadores sin signos. 8 Los brotes
de coriza infecciosa ocurren frecuentemente en otoo
e invierno. 9 No se ha demostrado que los gorriones
silvestres (Paser paser) estn implicados como vectores;
sin embargo, estudios epidemiolgicos sugieren que
este microorganismo puede ser introducido en granjas
aisladas por va area.10

Periodo de incubacin
El periodo de incubacin de la coriza infecciosa es de
24 a 48 h despus de la inoculacin de aves con cultivo
vivo o exudado infeccioso. De manera experimental,
el periodo de incubacin puede ser variable de
acuerdo con ciertas condiciones de exposicin: 24 h,
inoculacin intrasinusal; 48 h, instilacin nasal; 72 h,
aves en jaula; cuatro das, contacto con agua infectada
y seis a 14 das por transmisin area.11

Signs

Signos

Characteristic signs of infectious coryza include serous

or mucous nasal exudates, sneezing, swelling of
infraorbital sinuses, facial edema, and conjunctivitis.1
The swelling of wattles is mainly evident in males
(Figure 1). Also, tracheal rales may be listened when
the lower respiratory tract is affected.
In some cases, when H. paragallinarum causes
respiratory disease together with other infectious
agents, a severe clinical outcome may be produced.
The most common agents are: Mycoplasma sinoviae,12
M. gallisepticum,13 Ornithobacterium rhinotracheale,14,15
Escherichia coli, Salmonella spp, Pasteurella spp and
infectious bronchitis virus,16,17 among others.
Malkinson et al.18 reported cases of H. gallinarum
associated with Chlamydia psittaci and smallpox virus in
broiler breeders. However, association with Pasteurella
gallinarum may be frequently found, a bacterium that
may appear after the acute phase of infectious coryza,
causing a severe purulent panoftalmy and caseous
material contents into the parasanal sinuses.17
Chickens may have diarrhea, and feed and water
consumption are usually decreased. A bad growing
performance in broiler flocks may be registered. More
commonly found is a marked egg production drop in
laying hen flocks; in very severe cases a decreased of
up to 58.7% have been reported.19

Los signos caractersticos de la coriza infecciosa

incluyen exudado nasal seroso o mucoso, estornudo,
inflamacin de senos infraorbiarios, edema facial
y conjuntivitis.1 La inflamacin de barbillas puede
ser particularmente evidente en machos (Figura 1).
Tambin se puede escuchar estertor traqueal cuando
las aves tienen afectado el tracto respiratorio inferior.
Se ha observado un cuadro respiratorio ms severo
en casos donde se asocia H. paragallinarum con
otros agentes: Mycoplasma sinoviae,12 M. gallisepticum,13
Ornithobacterium rhinotracheale,14,15 Escherichia coli,
Salmonella spp, Pasteurella spp y virus de la bronquitis
infecciosa,16,17 entre otros. Malkinson et al.18
informaron la asociacin de H. gallinarum con
Chlamydia psittaci y el virus de la viruela aviar en
reproductores pesados. Sin embargo, parece muy
comn la asociacin con Pasteurella gallinarum, bacteria
que puede aparecer luego de la fase aguda de la coriza
infecciosa y causa panoftalma purulenta y contenido
de masas caseosas en los senos paranasales.17
Las aves pueden tener diarrea y el consumo de
agua y alimento generalmente se reduce. En aves en
crecimiento se registra mala utilidad de la parvada;
en gallinas de postura la reduccin en la produccin
de huevo puede llegar a 58.7%.19

Morbilidad y mortalidad
Morbidity and mortality
Clinically, classic infectious coryza is usually
characterized by high morbidity and low mortality
rates.1 In addition, non-classic clinical outbreaks with
high mortality caused by H. paragallinarum per se have
also been reported. Losses due to persistent mortality
and culling were up to 2%-5% in both growing
and layer flocks.17 In these cases, H. paragallinarum
was isolated from liver, kidney, and particularly from
tarsus and internal contents of eyeballs indicating a

La coriza infecciosa clsica est generalmente

caracterizada por alta morbilidad y baja mortalidad.1
Sin embargo, se ha informado de cuadros clnicos
atpicos donde H. paragallinarum per se ha causado
mortalidad. En parvadas de pollos de engorda y
gallinas de postura, las prdidas debidas a mortalidad
persistente y eliminacin de aves fue de 2%-5%.17 En
estos casos, H. paragallinarum fue aislado a partir de
hgado, rin y especialmente de la articulacin del
tarso y globos oculares, ell indica septicemia. Droual

Figura 1. Pollo infectado artificialmente con Haemophilus paragallinarum. Se observa coriza, inflamacin de seno infraorbitario
y edema de barbilla
Artificially infected chicken Haemophilus paragallinarum. Coryza,
swelling of infraorbital sinus, and wattle edema are observed.

Vet. Mx., 35 (3) 2004

263

septicemia. Droual et al.20 described condemnations

of broilers in the slaughterhouse, mainly due to
fibrinopurulent cellulitis lesions of the head and
wattles together with airsacculitis.
Bland et al. 21 informed the most severe infectious
coryza outbreak recorded until now in a laying
complex located the California area. During a period
of 13 weeks, mortality varied from 8% up to 64% and
egg production dropped from an average of 77% to
only 15%.

et al.20 describieron descartes en matadero de pollos

de engorda por lesiones de celulitis fibrinopurulenta
de la cabeza y barbillas y aerosaculitis. Bland et al.21
informaron que en un complejo de gallinas de postura
en California, durante un periodo de 13 semanas, la
mortalidad vari del 8% al 64% y la produccin de
huevo cay de un promedio de 77% a tan slo 15%, en
el brote de coriza infecciosa ms severo registrado a la
fecha en esta rea.

Hallazgos macroscpicos y microscpicos

Macroscopic and microscopic findings

Chickens affected with H. paragallinarum have serous or
mucous exudates in their nasal cavity and infraorbital
sinuses. 22 In the mucosal membranes of these tissues,
hydropic or edematous inflammation (acute catarrhal
inflammation) is observed. Also, subcutaneous edema
in the periorbital area and wattles is observed (Figure
2). 23 Microscopic changes are limited to mucosal
surfaces of nasal passages and infraorbital sinuses.
The primary lesion consist of a citotoxic effect in the
epithelium and a marked stimulation of intraepithelial
globet cells activity. 24 Also, a marked mast cell
infiltration into the mucosal membranes of the nasal
cavity is observed. The secreted products from mast
cells, heterophils and macrophages are responsible for
the severe vascular changes and cellular damage that
lead to coryza. 23 However, it is still not well understood
the pathogenesis of H. paragallinarum in the infectious
coryza.

Molecular epizootiology
In addition to serotyping and biotyping, Blackall et
al.8 included restriction endonucleases to study 16
cases of infectious coryza from northern New South
Wales, Australia. These endonuclease patterns allowed

En pollos infectados con H. paragallinarum, la cavidad

nasal y senos infraorbitarios presentan exudado seroso
o mucoso. 22 Las membranas mucosas de estos sitios se
observan congestionados con inflamacin hidrpica
o edematosa (inflamacin catarral aguda). Tambin
se observa edema en el tejido subcutneo de la
regin periorbital y de las barbillas. 23 Los cambios
microscpicos se limitan principalmente a la mucosa
de los pasajes nasales y senos infraorbitarios. La lesin
bsica observada es un infiltrado inflamatorio de
la mucosa respiratoria con indicacin de un efecto
citotxico en el epitelio y marcada estimulacin de
las glndulas mucosas intraepiteliales. 24 Tambin se
observa infiltracin marcada de mastocitos en la
lmina propia de las membranas mucosas de la
cavidad nasal. Los productos de los mastocitos,
heterfilos y macrfagos pueden ser responsables de
los cambios vasculares severos y el dao celular que
conduce a coriza. 23 Sin embargo, an no se comprende
bien la patogenia de H. paragallinarum en la coriza
infecciosa.

Epizootiologa molecular
De forma adicional a pruebas de serotipificacin
y biotipificacin, Blackall et al.8 incluyeron

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264

Figura 2. Descripcin del procedimiento para la

obtencin de Haemophilus paragallinarum a partir de
senos infraorbitarios de aves con coriza infecciosa.
Detailed Haemophilus paragallinarum isolation procedure from an infraorbital sinus of infectious coryza diseased chicken.

the recognition of three groups of related outbreaks.

These results showed that the farms may have a chronic
infection caused by an unique H. paragallinarum
strain, which intermittently appears. This study also
showed that replacement pullets are the main source
of infectious coryza.
Similarly Miflin et al.,25 characterized a total of
15 NAD-independent H. paragallinarum, isolated since
1989 in South Africa by means of serotyping, biotyping,
restriction endonuclease analysis and rybotyping.
All isolates were serovar A and showed an unique
restriction endonuclease and rybotyping pattern,
which was different from the ones obtained when
NAD-dependent isolates were analyzed before 1989.
Based on these results, the authors suggested that
the NAD-independent isolates belong to a classical
strain clon. Similarly, the molecular epidemiological
rybotyping was performed for 12 serovar A H.
paragallinarum strains, isolated from five infectious
coryza outbreaks in the Chinese Province of Hebei. 26
The results showed four rybotyping patterns, which
were matched to the epidemiological history of these
isolates, confirming the existence of H. paragallinarum
genetic diversity in China.
A total of 15 Blackall serovar isolates of H.
paragallinarum were studied by means of the ERICPCR technique in Mexico by Soriano and Blackall. 27
Two ERIC profiles were showed for the serovar A-1
isolates; one for the serovar A-2 isolate; three for
serovar B-1 isolates, and two for the serovar C-2 isolates.
A relationship between the serovar and the ERIC
profile was established based on the origin of isolates.
The authors concluded that the ERIC-PCR technique
could be a rapid laboratory tool for epidemiological
characterization of H. paragallinarum.

Pathogenicity and virulence

As previously mentioned, the hemagglutinating
antigens, or hemagglutinins, are structures that are
mainly related to the antigenicity, pathogenicity, and
immunogenicity of H. paragallinarum. In this way, a
non-hemagglutinating variant strain, which is unable
to hemagglutinate (even after laboratory treatments
or aging) is also unable to produce infectious coryza
after instillation or inoculation of susceptible birds. 28,29
Takagi et al.30 purified this hemagglutinin, using
affinity chromatography and monoclonal antibodies,
and demonstrated its crucial immunogenic role
when they achieved protection in susceptible birds
challenged with a pathogenic strain by previously
immunizing the birds with the purified hemagglutinin
by intramuscular route. Furthermore, this active
protection was completely dependent on the
hemagglutination-inhibition antibodies present in

endonucleasas de restriccin en el estudio

epidemiolgico de 16 casos de coriza infecciosa en
el norte de New South Wales, Australia. El perfil
obtenido permiti reconocer tres grupos de brotes
relacionados. Los resultados indicaron que las granjas
pueden tener infeccin crnica causada por una sola
cepa de H. paragallinarum que reaparece a intervalos.
Este estudio tambin mostr la primera evidencia
detallada de que las aves de reemplazo son la mayor
fuente de coriza infecciosa.
De forma similar, Miflin et al.25 caracterizaron
15 aislamientos de H. paragallinarum independientes
de NAD obtenidos a partir de 1989 en Sudfrica,
mediante serotipificacin, biotipificacin, anlisis con
endonucleasas de restriccin y ribotipificacin. Todos
los aislamientos fueron serovariedad A y mostraron
un nico patrn en el anlisis con endonuclesas
de restriccin y ribotipificacin, que fue diferente a
los patrones de aislamientos dependientes de NAD
obtenidos antes de 1989. Basados en lo anterior, los
autores sugirieron que los aislamientos independientes
de NAD corresponden a clones de las cepas clsicas.
En un estudio similar, mediante perfiles de ribotipificacin se estableci la epidemiologa molecular de
12 aislamientos (serovariedad A) de H. paragallinarum
obtenidos en cinco brotes de coriza infecciosa en
Hebei, China. 26 Los resultados obtenidos mostraron
cuatro perfiles de ribotipificacin, los cuales se
correspondieron con la historia epidemiolgica de
los aislamientos, confirmando que existe diversidad
gentica en la poblacin de H. paragallinarum de ese
pas.
En Mxico, Soriano y Blackall, 27 mediante la
prueba de ERIC-PCR estudiaron 15 aislamientos de
H. paragallinarum de las serovariedades de Blackall.
Los resultados mostraron dos perfiles ERIC en los
aislamientos serovariedad A-1, uno para el aislamiento
A-2, tres para los aislamientos B-1 y dos para los
aislamientos C-2. Con base en el origen de los
aislamientos, los autores establecieron una relacin
entre la serovariedad y el perfil ERIC obtenido,
concluyeron que esta prueba puede ser una
herramienta de laboratorio rpida en la
caracterizacin epidemiolgica de H. paragallinarum.

Patogenicidad y virulencia
Los antgenos hemoaglutinantes, o hemoaglutininas,
son las estructuras principalmente relacionadas con
la antigenicidad, patogenicidad e inmunogenicidad
de H. paragallinarum. As, una cepa variante que
no hemoaglutina, aun despus de tratamientos o
envejecimiento, tampoco produce coriza cuando se
instila o inocula en aves susceptibles. 28,29 Takagi
et al.30 purificaron esta hemoaglutina mediante el

Vet. Mx., 35 (3) 2004

265

the blood serum. Takagi et al.31,32 demonstrated

a solid passive protection against challenges with
the homologous strain when specificaly against,
mouse monoclonal antibodies hemagglutinin were
administered into the peritoneal cavity of chickens.
Despite all these studies were carried out using 221
(A-1) strain , it is very likely that these results could be
extended and applied to the hemagglutinins of most
H. paragallinarum strains.
Bacterial adherence to epithelial cells is regarded
as the first step of infection of the mucosal surfaces.
Adhesins are structures that mediates adhesion to the
complementary cellular structures, the receptors. 33
Both in vivo and in vitro positive adherence of H.
paragallinarum to chicken tracheal epithelial cells have
been shown. 34,35 The treatment by an homologous or
heterologous adsorption of H. paragallinarum bacterial
cells, either with rabbit or chicken antisera or with
tracheal washings from immunized chickens, produce
the complete loss of the hemagglutinating activity
and adherence ability of this bacterium to chicken
epithelial cells. 36 These results showed that the
hemagglutinin antigens truly are the adhesins of H.
paragallinarum and the hemagglutination-inhibition
antibodies that exist in the tracheal washings really act
as neutralizing antibodies. This fact indicates that the
specific hemagglutinin receptors located in both the
erythrocytes and the respiratory epithelial cells are
the same, or very similar, confirming the importance
of these hemagglutination tests which indirectly
detect the pathogenic ability of a particular strain
of H. paragallinarum to adhere to epithelial target
cells. Following the same methodology it was showed
that the main cellular receptor may be related
to the carbohydrate D-mannose. 37 Among several
Pasteurellaceae members, this carbohydrate have also
been identified as the main cellular receptor. 31
Recently, Terry et al.38 reported haemocin
production, a bacteriocin, from H. paragallinarum.
Furthermore, a chromosomal gene encoding for this
haemocin, as well as a plasmid carrying this gene
in an Australian isolate was identified. As pointed
out by these authors, haemocin production by H.
paragallinarum could be an important colonization
factor of the chicken respiratory sinuses. They showed
that isolates of Pasteurella avium, P. volantium and P.
species A, are all of them regarded as non-pathogenic
bacteria found in the respiratory tract of chickens,
were all sensitive to this haemocin.
H. paragallinarum is regarded as a primary
pathogenic agent for susceptible chickens.1 However,
specific discrepancies about the pathogenicity of the
0222 (serovar B-1) strain have subsisted. A lack of
agglutinin antigens and a non-pathogenic outcome
in challenged chickens were reported by Sawata et

266

uso de cromatografa de afinidad y anticuerpos

monoclonales y demostraron su importancia crucial en
la inmunogenicidad al inocular con la hemoaglutinina
purificada aves susceptibles por va intramuscular. De
este modo se logr proteccin frente a desafos con
una cepa patgena. Es ms, esta proteccin activa
depende totalmente de la presencia de anticuerpos
humorales inhibidores de la hemoaglutinacin en el
suero sanguneo. Adems, Takagi et al.31,32 demostraron
que se puede conferir una slida proteccin frente
al desafo con la misma cepa, administrando por va
intraperitoneal anticuerpos monoclonales de ratn,
especficamente dirigidos contra las hemoaglutininas;
esta vez, mediante un mecanismo de inmunidad
pasiva. Si bien estos ltimos estudios se realizaron con
la cepa 221 (A-1), estos resultados son, con certeza,
generales y por extensin pueden ser aplicados a las
hemoaglutininas de la mayora de las cepas de H.
paragallinarum.
Se considera que la adherencia bacteriana a
las clulas epiteliales es el primer paso en el
proceso de infeccin de las superficies mucosas. Las
adhesinas son las estructuras bacterianas que median
la adherencia a estructuras celulares complementarias,
los receptores. 33 Con base en lo anterior, se ha
mostrado adherencia in vivo e in vitro de H.
paragallinarum a clulas epiteliales traqueales de
pollo. 34,35 De forma similar, se ha mostrado que
bacterias de H. paragallinarum, adsorbidas de manera
homloga o heterloga con antisueros de conejo
o pollo y lavados traqueales de aves inmunizadas,
perdieron la actividad hemoaglutinante y capacidad
de adherencia a las clulas epiteliales. 36 Los resultados
obtenidos en este estudio indican que los antgenos
hemoaglutinantes son las adhesinas de H.
paragallinarum y que los anticuerpos inhibidores de la
hemoaglutinacin presentes en los lavados traqueales
actan tambin como anticuerpos neutralizantes.
Este hecho indica que seguramente los receptores
especficos para las hemoaglutininas de los eritrocitos
y las clulas epiteliales del tracto respiratorio son los
mismos, o muy similares, lo cual una vez ms confirma
la importancia de las pruebas de hemoaglutinacin, las
cuales, indirectamente, detectan la capacidad de una
determinada cepa para adherirse a las clulas blanco
sobre las que H. paragallinarum especficamente dirige
su accin patgena. Siguiendo esta metodologa,
tambin se mostr que el receptor celular puede
ser principalmente del tipo D-manosa. 37 Se ha
comprobado que este carbohidrato es tambin el
receptor celular para algunos miembros de la familia
Pasteurellaceae. 31
Recientemente, Terry et al.38 informaron de la
produccin de hemocina, una bacteriocina, por parte
de H. paragallinarum. Adicionalmente identificaron el

al. 39 Conversely, Rimler et al.40,41 showed that 0222

and Spross (B-1) strains were both pathogenic in
challenged chickens, representing one of the three
serovar-related, immunovars identified by the slide
agglutination tests. Similarly, Thornton and Blackall42
found that 0222 strain produced serovar-specific,
agglutination antibodies. The pathogenicity of five
serovar B strains that included strains 0222 and Spross,
was investigated by Yamaguchi et al. 43 Contrasting
to the pathogenicity observed with the other three
strains, they found that 0222 strain did not produce
any clinical signs of infectious coryza. Nevertheless,
lesions in the infraorbital sinuses (thickness of mucosal
membranes with yellowish exudate) and isolation of
the challenged microorganism were observed. These
results show that 0222 strain is able to produce
infection and also point out that it is probably a low
virulent strain. However, definitive studies have not
been carried out.
Bragg et al.44 recorded several virulence degrees
in chickens challenged with South African isolates
of serovars A-1, B-1, C-2, and C-3. The most severe
disease signs in challenged chickens were produced
by the serovar C-3. In a similar way, Soriano et al.45
independently challenged chickens with each of the
nine Blackalls serovar reference strains and showed
that the most severe clinical signs of infectious coryza
were observed when the chickens were inoculated
with the serovars C-1 and C-3.
Sandoval and Terzolo 29 developed different
experimental disease models to study the
pathogenicity, horizontal spread and invasiveness of
several isolates from Argentina, belonging to serovars
A, B, and C. They showed that the serovar B strains
were consistently very pathogenic causing serious
acute coryza lesions with a high degree of spread
and invasiveness. On the other hand, serovar A and
C strains showed a different pattern: some of them
were very virulent, spreading and invasive; others were
pathogenic but diffuse slowly; some strains showed
low pathogenicity and even a few field strains were
found to be completely non-pathogenic.
Horner et al.46 suggested that NAD-independent
isolates may more frequently cause airsacculitis than
typical H. paragallinarum isolates. Furthermore, it is
thought that these field NAD-independent isolates
are so different from typical strains included in the
standard vaccines that they may produce failures. 47
Virulence of NAD-independent isolates of H.
paragallinarum has been also investigated. Bragg48
challenged groups of chickens with NAD-independent
isolates and evidenced its pathogenicity. However,
the clinical signs observed in these chickens were
less severe than the ones produced when chickens
were challenged with NAD-dependent strains. It

gen cromosmico que codifica para esta hemocina, as

como un plsmido que porta este gen un aislamiento
de Australia. Los autores mencionan que la hemocina
producida por H. paragallinarum puede ser importante
en la colonizacin de los senos respiratorios de
los pollos, ya que encontraron que aislamientos de
Pasteurella avium, P. volantium y P. especie A, todos ellos
considerados bacterias gramnegativas no patgenas y
que se encuentran en el tracto respiratorio superior
de pollos que sufren enfermedad respiratoria debida
a otros agentes, fueron sensibles a esta hemocina.
Se considera que H. paragallinarum es un agente
patgeno primario en aves susceptibles.1 Sin embargo,
de manera especfica han existido discrepancias
en cuanto a la patogenicidad de la cepa 0222
(serovariedad B-1). Sawata et al. 39 informaron que
esta cepa careca de antgenos aglutinantes y que
no era patgena en pollos desafiados. Sin embargo,
Rimler et al.40,41 mostraron que las cepas 0222 y
Spross (B-1) fueron patgenas en pollos desafiados y
que representaba una de las tres inmunovariedades
relacionadas con las serovariedades identificadas por
pruebas de aglutinacin en placa. De forma similar,
Thornton y Blackall42 encontraron que la cepa 0222
haba producido anticuerpos aglutinantes especficos
de serovariedad. Yamaguchi et al.43 investigaron la
patogenicidad de cinco cepas serovariedad B, las
cuales incluyeron las cepas 0222 y
Spross. A
diferencia de las otras tres cepas, la cepa 0222
no produjo signos clnicos de coriza infecciosa. Sin
embargo, se observaron lesiones (engrosamiento de
membranas mucosas y exudado amarillento) en senos
infraorbitarios y aislamiento del microorganismo de
desafo. Estos resultados evidencian que la cepa 0222 es
capaz de producir infeccin e indican, probablemente,
que es una cepa de baja virulencia. Sin embargo, no se
han conducido los estudios definitivos al respecto.
Bragg et al. 44 registraron diferentes grados de
virulencia en pollos desafiados con aislamientos
sudafricanos de las serovariedades A-1, B-1, C-2 y C-3.
El aislamiento serovariedad C-3 produjo los signos
ms graves en las aves desafiadas. De forma similar,
Soriano et al.45 desafiaron grupos de pollos de manera
independiente con las nueve cepas de referencia del
esquema de Blackall, e informaron que los signos
ms graves de coriza infecciosa se observaron en aves
desafiadas con las cepas de las serovariedades C-1 y
C-3.
Sandoval y Terzolo 29 estudiaron diferentes cepas
regionales de las serovariedades A, B y C de Argentina,
mediante el desarrollo de modelos de reproduccin
de la enfermedad para evaluar su patogenicidad,
difusin horizontal e invasividad. Se demostr que las
cepas B fueron consistentemente patgenas y causan
lesiones de coriza muy aguda, siempre con alto grado

Vet. Mx., 35 (3) 2004

267

is important to point out that NAD-independency

could be acquired by transformation by means of
a plasmid, as reported by Bragg et al.49 Similarly, a
NAD-dependent C-3 strain was transformed into a
NAD-independent strain by means of electroporation
technique carried out by Taole et al.50 Less severe
infectious coryza signs were observed when chickens
were challenged with the transformed strain. The
mechanism by which virulence is affected in this
process was not explained. An interesting practical
application of this transformation could be a possible
reduction in the vaccine production costs by the use
of NAD-independent strains in industrial cultures.
In fact, as was demonstrated by Bragg et al.,51
these transformed strains maintain unchangeable
the production of hemagglutinins, basic antigens
to achieve a good vaccine protection. However, the
introduction of infective plasmids into countries
free from NAD-independent strains arise serious
biosecurity problems, particularly taking into account
that in South Africa also exist NAD-independent
strains of P. avium, P. volantium and Pasteurella species
A in chickens, 47 and even in H. parainfluenzae strains
isolated from human beings, 52 indicating that in this
country it may have occurred a widespread infection
of this plasmid. 53
It has been reported that the capsule of H.
paragallinarum could be involved in pathogenicity
and virulence of this bacterium. 54-57 Nevertheless,
definitive studies showing the role of the capsule in
the pathogenesis of H. paragallinarum have not been
carried out.
Increased gallinacin-3 expression in experimentally
H. paragallinarum infected chickens was recently
reported. 58 Gallinacin-3 is an epithelial -defensin
involved in the innate immunity by means of
antimicrobial properties of the epithelial cells and
tracheal secretions. It could be a limiting factor for
tissue dispersion of H. paragallinarum in the respiratory
tract and for the colonization processes in chickens.

Diagnosis
Isolation and identification of the agent
Three to five birds with acute coryza signs are
recommended for bacteriological isolation. In
addition, strict sterility is recommended for sampling.
Once the bird has been euthanatized, the skin of the
infraorbital area is cauterized and an incision is made
onto the corresponding infraorbital sinus. The skin is
separated and a sterile swab is introduced, previously
moistened into nutritive broth or phosphate buffer
solution at neutral pH (Figure 2). Due to the reduced
viability of H. paragallinarum, it is recommended

268

de contagio e invasividad. En cambio las cepas A y C

mostraron distinto comportamiento: algunas fueron
muy virulentas, difusoras e invasivas; otras fueron
patgenas, pero slo difundan muy lentamente, y
otras fueron poco patgenas e inclusive se encontraron
unas pocas cepas de campo que mostraron ser
totalmente apatgenas.
Horner et al.46 sugirieron que los aislamientos
independientes de NAD pueden causar aerosaculitis
de manera ms frecuente que los H. paragallinarum
tpicos. Adems, se especula que los aislamientos
independientes de NAD pueden ser diferentes como
para producir fallas de las bacterinas actualmente
empleadas. 47
La virulencia de aislamientos de H. paragallinarum
independientes de NAD tambin ha sido investigada.
Bragg48 desafi grupos de aves con aislamientos
independientes de NAD en los cuales su patogenicidad
fue evidente. Sin embargo, los signos de coriza
infecciosa fueron menores que los observados en
aves desafiadas con cepas dependientes de NAD. Es
importante destacar que Bragg et al.49 han demostrado
que la independencia de NAD puede ser adquirida
por transformacin mediante la adquisicin de un
plsmido. De forma similar, Taole et al. 50 transformaron
una cepa serovariedad C-3 dependiente de NAD en
independiente de este factor, mediante la tcnica
de electroporacin. Los signos de coriza infecciosa
fueron menos graves en las aves que fueron desafiadas
con la cepa transformada. No se pudo explicar el
mecanismo por el cual la virulencia es afectada por
este proceso. Lo interesante de esta transformacin
es su posible aplicacin prctica, ya que sera factible
reducir el costo de produccin de las bacterinas
al usar cepas que no requieren de la adicin de
NAD en los cultivos industriales. De hecho, Bragg
et al. 51 demostraron que estas cepas trasformadas
mantienen inalterables su capacidad de producir
hemoaglutininas, antgenos fundamentales para
lograr una buena proteccin en la bacterinizacin.
Sin embargo, la posible introduccin de plsmidos
infectivos en pases carentes de cepas independientes
de NAD, plantea muy serios problemas de
bioseguridad, sobre todo si se toma en cuenta que en
Sudfrica, por ejemplo, adems de H. paragallinarum,
tambin existen cepas NAD independientes de P.
avium, P. volantium y Pasteurella especie A en las aves, 47
e inclusive de H. parainfluenzae en seres humanos, 52 lo
que indica que podra existir una posible infeccin
por este plsmido. 53
Se ha informado que la cpsula de H. paragallinarum
est implicada en la patogenicidad y virulencia de
esta bacteria. 54-57 Sin embargo, no se han efectuado
estudios definitivos que muestren la importancia de la
cpsula en la patogenia de H. paragallinarum.

to perform the culture within five hours of the

euthanasia. Both agar base or Columbia agar base
plates plus 7% bovine or sheep blood with the addition
of Staphylococcus spp as nurse colony could be used
for culturing the swabs. Chocolate agar or any
of the aforementioned base agar plus haemolyzed
blood may be used instead of the nurse colony. As
we have previously mentioned, bigger colonies are
obtained using this method. A procedure that allows
differentiation and isolation of pure H. paragallinarum
cultures, even when the accompanying bacterial flora
is complex, is the use of a selective media with
added antibiotics and incubated at 37C for 48 h into
a microaerophilic atmosphere. The microaerophilic
atmosphere may be obtained by means of the classical
candle jar, consuming part of oxygen contained into a
hermetic closed container, or by the usage of different
available commercial products, to generate either
CO 2 or the commercial atmospheres for isolating
bacteria of the Campylobacter genus. 53 Biochemical
and enzymatic tests have to be performed to identify
H. paragallinarum. NAD-dependency or independency
does not allow the differentiation of H. paragallinarum
from other microorganisms of the Pasteurella genus,
particularly in places where NAD-independent
bacteria also exist. 59 Up to now, NAD-independent
strains of H. paragallinarum from South Africa (serovars
A-1 and C-3) 60,61 and from Mexico (serovars B-1 and
C-2) have only been reported. 62

Serological identification
A number of serological tests have been reported
for the detection of antibodies in chickens against
H. paragallinarum : gel precipitation, 63 slide
agglutination, 64 latex agglutination 65 and ELISA. 66-68
However the hemagglutination-inhibition test is the
most widely used. 69
Several monoclonal antibody panels have been
used for the identification of H. paragallinarum, mainly
by hemagglutination-inhibition and ELISA tests.70-74
Monoclonal antibodies that recognized 49 serovar
A isolates from Japan77 and more than 20 serovar
A71,72 isolates from several parts of the world, failed
to recognize 4 out of 10 serovar A75 isolates from
Argentina and 6 out of 14 serovar A76 isolates from
Brazil, showing that these Latin American strains have
certain antigenic differences.

Molecular identification
A specific polymerase chain reaction (PCR) for H.
paragallinarum was developed by Chen et al.78 This
is a rapid test and results could be obtained in
approximately 6 h. Also, both NAD dependent or

Un estudio reciente mostr que pollos infectados

de manera experimental con H. paragallinarum,
incrementaron la expresin de gallinacina-3, una
-defensina epitelial que contribuye a la inmunidad
innata mediante propiedades antimicrobianas propias
de las clulas epiteliales y secreciones traqueales. 58
Los autores mencionan que es probable que ste
sea un factor limitante de la distribucin tisular
de H. paragallinarum en el proceso de infeccin y
colonizacin del tracto respiratorio de los pollos.

Diagnstico
Aislamiento e identificacin del agente
Para el aislamiento bacteriolgico se recomienda el
estudio de tres o cinco aves con signos agudos de
coriza. El procedimiento de toma de muestras se debe
efectuar con estricta esterilidad. Para ello, una vez
sacrificada el ave, se cauteriza la piel de la regin
infraorbital y se practica una incisin sobre el seno
infraorbitario correspondiente, se separa la piel en la
incisin y se introduce un hisopo estril humedecido
en un caldo nutritivo o solucin tamponada de fosfatos
a pH neutro (Figura 2). Lo ms recomendable es el
cultivo antes de las 5 h debido a la reducida viabilidad
de H. paragallinarum. Para la siembra de los hisopos
pueden utilizarse placas en base de agar, o agar
Columbia con 7% de sangre de bovino u ovino con
el agregado de cepas nodriza de Staphylococcus spp, las
cuales eliminan el factor V, o bien usar agar chocolate
o agar con sangre hemolizada, en vez de las cepas
nodriza. Como se refiri anteriormente, con este
ltimo procedimiento se obtienen colonias mucho
ms grandes. El uso de medios de cultivo selectivos
con antibiticos e incubados a 37C durante 48 h en
una atmsfera microaeroflica, es un procedimiento
que permite diferenciar y aislar a H. paragallinarum
en cultivo puro, aun cuando la flora bacteriana
sea compleja. La atmsfera microaeroflica puede
obtenerse mediante el clsico mtodo de incubacin
de las placas en un recipiente con vela, la cual se
apaga al consumirse parte del oxgeno contenido en
un recipiente hermticamente cerrado, o bien usando
los distintos productos comerciales disponibles, tanto
para generar CO 2 como para las atmsferas destinadas
al gnero Campylobacter. 53 La identificacin de H.
paragallinarum debe efectuarse mediante pruebas
bioqumicas y enzimticas diferenciales. La
dependencia o independencia de NAD no permite
inferir si se trata de H. paragallinarum o de otros
microorganismos del gnero Pasteurella, sobre todo
en los lugares donde existen cepas independientes de
NAD. 59 A la fecha, nicamente se ha informado el
aislamiento de H. paragallinarum independientes de

Vet. Mx., 35 (3) 2004

269

independent isolates can be identified.79 This test is

called HP-2 PCR and use the following primers that
amplify a 0.5 kb fragment78: N1, 5 TGA GGG TAG
TCT TGC ACG CGA AT 3 and R1, 5 CAA GGT
ATC GAT CGT CTC TCT ACT 3. This technique
was developed in Australia and have been successfully
transferred to South Africa,79 China 80 and Mexico. 81
Samples directly obtained from the infraorbital sinuses
of experimentally infected chickens have shown
excellent results in further studies.78 Similarly, positive
results are obtained when samples are stored at 4 or
20C for 180 days. 82

Molecular characterization
The restriction endonucleases BamHI, EcoRI HindIII
and SmaI were evaluated for DNA characterization of
H. paragallinarum by Blackall et al.83 More fragments
and discrimination ability among the studied isolates
were obtained when the enzyme HindIII was used.
The PCR development for H. paragallinarum was
based on the creation of a genomic library for Modesto
(serovar C-2) strain.78 Based on the study of this
genomic library, four probes that specifically reacted
with H. paragallinarum were identified. None of these
probes reacted neither with bacteria of the Pasteurella
and Actinobacillus genus, nor with M. gallisepticum and
M. synoviae. PCR primers were designed by using the
partial sequence of the smallest probe (P601, 1.8 kb).
An enterobacterial repetitive intergenic consensusbased PCR (ERIC-PCR) that identified 18 ERIC
patterns among 39 isolates and reference strains of
H. paragallinarum was reported by Khan et al. 84 With
exception of the Australian HP14 (A-4) and HP60
(C-4) reference strains that shared the same pattern,
each isolate or reference strain gave an unique ERIC
pattern. However, in a similar study that included both
aforementioned Australian reference strains, Soriano
and Blackall 27 found differences in the ERIC patterns
of these two strains.
Recently, the gene encoding for a hemagglutinin of
H. paragallinarum, HagA, was identified and sequenced
by Hobb et al.85 A little variation of the sequence of
11 reference strains was shown. A higher variation
in the amino acids sequence was expected because
the hemagglutinin is the main antigen involved in
serotyping. However, no correlation was observed
between these variations and the serological groups of
the strains. As will be mentioned afterwards, probably
the HagA gene encodes for a common hemagglutinin
identified in H. paragallinarum strains.

Differential diagnosis
A confident diagnosis of infectious coryza is being

270

NAD en Sudfrica (serovariedades A-1 y C-3) 60,61 y

Mxico (serovariedades B-1 y C-2). 62

Identificacin serolgica
Se han descrito varias pruebas serolgicas para la
deteccin de anticuerpos contra H. paragallinarum
en los pollos: precipitacin en gel, 63 aglutinacin
en placa, 64 aglutinacin en ltex 65 y ELISA. 66-68
Sin embargo, la prueba de inhibicin de la
hemoaglutinacin es la ms usada. 69
Se han producido un nmero de paneles de anticuerpos monoclonales que han sido empleados para
identificar H. paragallinarum, principalmente mediante
pruebas de inhibicin de la hemoaglutinacin y
ELISA.70-74 En Argentina, cuatro de diez aislamientos
de la serovariedad A75 y en Brasil seis de 14 aislamientos de esta misma serovariedad76 no reaccionaron
con los anticuerpos monoclonales especficos que
reconocieron a 49 aislamientos japoneses de la serovariedad A77 y ms de 20 aislamientos de la serovariedad
A de varias partes del mundo; 71,72 en estas cepas
de Latinoamrica se observaron ciertas diferencias
antignicas.

Identificacin molecular
Chen et al.78 desarrollaron una prueba de reaccin
en cadena de la polimerasa (PCR, por sus siglas en
ingls, polymerase chain reaction), especfica para H.
paragallinarum. Esta prueba es rpida y los resultados
se obtienen aproximadamente en 6 h. Asimismo,
identifica aislamientos tanto dependientes como
independientes de NAD.79 Esta prueba es llamada
HP-2 PCR y utiliza los siguientes iniciadores: N1, 5
TGA GGG TAG TCT TGC ACG CGA AT 3 y R1,
5 CAA GGT ATC GAT CGT CTC TCT ACT 3, que
amplifican un fragmento de 0.5 kb.78 Esta prueba
fue desarrollada en Australia y ha sido transferida
con xito a Sudfrica,79 China 80 y Mxico. 81 Estudios
posteriores han mostrado excelentes resultados
cuando las muestras incluidas en la prueba son
tomadas directamente de senos infraorbitarios de
aves infectadas de manera experimental.78 De forma
similar, se obtienen resultados positivos en muestras
mantenidas a 4C o 20C durante 180 das. 82

Caracterizacin molecular
Blackall et al.83 evaluaron las endonucleasas de
restriccin BamHI, EcoRI HindIII y SmaI en la
caracterizacin de ADN cromosmico de H.
paragallinarum. La enzima HindIII produjo el mayor
nmero de fragmentos y mostr un mayor grado de
discriminacin entre los aislamientos estudiados.

more difficult because new bacteria or variants

have been identified in the recent years. One of
these microorganisms is O. rhinotracheale, bacterium
identified in Mexico and other countries.14,15 A coryzalike disease, characterized by growth retardation,
increased mortality and a significant drop in egg
production are produced by this Gram negative
bacterium. Airsacculitis and pneumonia are the
main observed lesions. In most of the countries,
differentiation of O. rhinotracheale and H.
paragallinarum it is not difficult. However, in places
like South Africa 40,60,61 and Mexico, 62 where NADindependent H. paragallinarum coexist together with
O. rhinotracheale, biochemical tests and carbohydrate
fermentation patterns are required to distinguish
both causal agents in coryza-like outbreaks. In
addition, NAD-dependent and independent strains
of P. volantium, P. avium and Pasteurella sp taxon A
are also involved in respiratory disease suggestive
of infectious coryza. 47 The swollen head syndrome
caused by turkey rhinotracheitis (TRT) virus in broiler
chickens have also to be differentiated. It has been
occasionally described that hypovitaminosis A and
chicken smallpox may produce coryza-like clinical
signs.7
Arthritis of tarsus caused by H. paragallinarum is
rarely found, but nevertheless in such cases differential
diagnosis from other bacteria or viruses is required. 86

Prevention and control

Immunity and vaccines
Vaccination of susceptible chicken flocks is the
most efficacious preventive practice against infectious
coryza. Experimental models using susceptible
chickens have been developed to evaluate crossprotection among different serovars and the protection
afforded by either experimental or commercial
vaccination programs. In these models the chickens
are first vaccinated with the commercial vaccines to
be tested and/or with monovalent vaccines prepared
with the same challenge strains to be studied
and then, after allowing a period for developing
active immunity, different flocks of vaccinated and
unvaccinated chickens are challenged by intrasinusal
or nasal instillation. Protection is regarded as the
percentage of chickens without disease, according to
an established criterion, and is evaluated at the second
or third day post-challenge. Evaluation is based on
a strict criterion, a chicken is regarded as diseased
if the following features are found: clinical signs of
infectious coryza, mucus in one or both infraorbital
sinuses, and isolation of H. paragallinarum in one or
two sinuses. 87,88 Regarding the clinical signs of coryza

La creacin de una genoteca de la cepa Modesto

(serovariedad C-2) sent las bases del desarrollo de la
prueba de PCR.78 El estudio de esta genoteca identific
cuatro sondas que reaccionaron de manera especfica
con H. paragallinarum. Ninguna de las sondas reaccion
con bacterias de los gneros Pasteurella y Actinobacillus,
o M. gallisepticum y M. synoviae. Se obtuvo la secuencia
de la sonda ms pequea (P601, 1.8 kb) que permiti
el diseo de los iniciadores para la prueba de PCR.
Basada en el empleo de esta tcnica para el consenso
intergnico repetitivo de enrobacterias (ERIC-PCR,
por sus siglas en ingls, enterobacterial repetitive intergenic
consensus), Khan et al.84 identificaron 18 patrones
ERIC en 39 aislamientos y cepas de referencia de
H. paragallinarum. Mencionan que todas las cepas de
referencia mostraron un patrn ERIC nico, excepto
las cepas HP14 (A-4) y HP60 (C-4), procedentes de
Australia. Sin embargo, en un estudio similar, Soriano
y Blackall 27 encontraron diferencias en los patrones
ERIC obtenidos para estas cepas.
Recientemente, Hobb et al.85 aislaron, identificaron
y obtuvieron la secuencia del gen HagA que codifica
para una hemoaglutinina de H. paragallinarum.
La secuencia de 11 cepas de referencia revel
un pequeo grado de variacin entre las cepas.
Como la hemoaglutinina es el principal antgeno de
serotipificacin, se esperaba encontrar variaciones en
la secuencia de aminocidos. Sin embargo, ninguna
de las variaciones se correlacion con los grupos
serolgicos de las cepas. Como se har mencin
posteriormente, es probable que el gen HagA codifique
para una hemoaglutinina comn identificada en cepas
de H. paragallinarum.

Diagnstico diferencial
En aos recientes se han identificado nuevas bacterias
o variantes en las aves, lo que hace ms difcil el
diagnstico confiable de coriza infecciosa. Uno de estos
microorganismos es la bacteria O. rhinotracheale, que
ha sido identificada en Mxico y otros pases.14,15 Esta
bacteria gramnegativa produce un cuadro corizoide
caracterizado por retraso del crecimiento, incremento
en la mortalidad y disminucin considerable de la
produccin de huevo. Las lesiones principalmente
observadas son aerosaculitis y neumona. La diferenciacin entre O. rhinotracheale y H. paragallinarum no es
difcil en la mayora de los pases. Sin embargo, en
Sudfrica 40,60,61 y Mxico, 62 donde estn presentes H.
paragallinarum independientes de NAD, se requieren
pruebas bioqumicas y patrones de fermentacin
de carbohidratos para establecer el agente causal
en cuadros corizoides. Otros microorganismos
implicados en pollos con enfermedad sugestiva de
coriza infecciosa son: P. volantium, tanto dependiente

Vet. Mx., 35 (3) 2004

271

como independiente de NAD, P. avium, tanto

dependiente como independiente de NAD y Pasteurella
sp taxon A, tanto dependiente como independiente
de NAD. 47 En pollos de engorda la coriza debera
ser diferenciada del sndrome de cabeza hinchada
por virus de la rinotraquetis infecciosa del pavo
(TRT). Tambin se cita que la hipovitaminosis A y la
viruela, en ocasiones, pueden producir signos clnicos
similares a coriza infecciosa.7
La artritis del tarso por H. paragallinarum es poco
frecuente, pero su descripcin seala la necesidad
de un diagnstico diferencial con otros agentes
bacterianos o vricos. 86

they are classified into four degrees of which degrees

2, 3, and 4 are considered to be coryza (positive)
in sensu estricto, whereas degrees 0 and 1 are both
considered to be negative (Figures 3 and 4).
It is widely accepted that the three Blackall
serogroups represent three different immunovars. 89
The accepted dogma is that there is no cross-protection
between serovars of these serogroups. Recently, the
cross-protection among all the nine reference strains
of H. paragallinarum of Blackalls scheme was evaluated
in vaccination/challenge trials by Soriano et al.45
These results confirmed that the three recognized
immunovars are different. However, partial crossprotection among serovars of the three serogroups
was observed. These results allow the obtainment
of preventive strategies against infectious coryza.
Nowadays bivalent (A-1 and C-1) and trivalent (A-1,
B-1, and C-2) infectious coryza vaccines are universally
used. Nevertheless up to now, some discrepancies
about serovar B protection conferred by bivalent
vaccines still subsist. 69,90 However, the need to use

Prevencin y control
Inmunidad e inmungenos
La bacterinizacin de parvadas susceptibles es la
estrategia ms eficaz en la prevencin de la coriza
infecciosa. Se han desarrollado modelos experimen-

Grados de enfermedad

1
Leve
Conjuntivitis
0

Grados de enfermedad

2
3

Positivos

272

Figura 3. Grados de coriza infecciosa. Interpretacin de

pruebas de proteccin al segundo da despus del desafo
de aves inmunizadas. Grado 0: Ave inoculada sin sntomas.
Considerada negativa. Grado 1: Leve conjuntivitis. Considerada negativa
Sickness degrees of infectious coryza. Regarded as negative.
Protection trial interpretation at second day post challenge
of vaccinated chickens. 0, challenged chicken with no clinical signs. 1, Slight conjunctivitis.
Figura 4. Grados de coriza infecciosa. Interpretacin de
pruebas de proteccin al segundo da despus del desafo de
aves inmunizadas. Grado 2: Conjuntivitis con el ojo parcialmente cerrado e hinchazn de zona periorbital y senos paranasales. Considerada positiva. Grado 3: Conjuntivitis con el
ojo totalmente cerrado, prpados no adheridos e hinchazn
notable de zona la periorbital y senos paranasales. Considerada positiva. Grado 4: Conjuntivitis con ojo totalmente
cerrado, prpados adheridos e hinchazn muy severa de
zona la periorbital y senos paranasales. Considerada positiva.
Degrees of infectious coryza. Regarded as positive.Protection
trial interpretation at second day post challenge of vaccinated chickens. 2, Partial-closed eye with conjunctivitis and
swelling of periorbital area and infraorbital sinus. 3, Conjunctivitis with totally closed eye, non adhered eyelids and
notorious swelling of periorbital area and paranasal sinus.
4, Conjunctivitis with totally closed aye, adhered eyelids and
severe swelling of periorbital area and paranasal sinus.

trivalent vaccines in areas where serovar B isolates

are present have been established by a number of
studies. For example, it has recently been reported
that a bivalent, reference strain-based vaccine does not
protected chickens challenged with a regional serovar
B-1 isolate in Mexico. 91 In other trials, Yamaguchi
et al.92 reported that bivalent vaccines conferred
good protection against Spross (B) strain, but failed
to protect the chickens when they were challenged
with regional South African B isolates. Furthermore,
Terzolo et al.88 demonstrated that even a trivalent
vaccine, which have included a reference serovar B
strain in the formulation, have failed to protect the
immunized chickens when they were challenged with
a local serovar B strain. In a comparative study,
Bowles et al.93 determined the genetic diversity and
relationships of 118 strains from six continents by
electrophorectical mobilization of eight metabolic,
chromosomal gene-encoded enzymes. It is interesting
to highlight that all the H. paragallinarum studied
strains were classified into two heterogeneous clusters,
while nine local, Argentinean serovar B strains,
occupied a different cluster. These results
demonstrated that the serovar B Argentinean strains
are genetically distant from any of the other studied
strains, regardless the serovar to which they belong.
It means that the Argentinean regional serovar B
strains are indeed quite different from any other
serovar B strains isolated from several parts of the
world. Recently, Jacobs et al.94 reported that trivalentcommercial vaccinated chickens were protected against
an Ecuadorian serovar C strain but not against serovar
B strains isolated from several countries (Ecuador,
Argentina, United States of America, and Zimbabwe).
Nevertheless all these serovar B isolates were wholly
recognized by antisera raised against Spross (B) and
H-18 (C-1) strains. Furthermore in these trials, serovar
B protection was increased when an Ecuadorian
serovar B strain was included in the trivalent vaccine.
Based on all these independent studies, serovar B
isolates (variants) may be consider to be different
from the standard B strains. Therefore characterized
variant B strains should be included in all
formulation of vaccines to be used in the geographical
areas in which B variant outbreaks occur. As up
to now all these variant B strains are serologically
indistinguishable from standard B reference strains,
they are not yet regarded as a new immunovar. Anyway,
the vaccine that included these variant B strains
has been provisionally designated as a tetravalent
vaccines. It is very likely that many of these reported B
isolates may constitute a new serovar (or serovars) into
the serogroup B, in a similar way as the serogroups A
or C include four serovars each. These results point out
the necessity of performing new serovar hemagglutinin

tales con aves susceptibles para evaluar la proteccin

cruzada entre distintas serovariedades, o bien el grado
de proteccin conferido por diferentes bacterinas
experimentales o comerciales combinadas en planes de
bacterinizacin. Estas pruebas consisten en inmunizar
a las aves con las bacterinas comerciales a probar
o bien con las bacterinas monovalentes elaboradas
con las cepas a estudiar y transcurrido un periodo
necesario para que las aves desarrollen inmunidad
activa, stas se desafan por va intrasinusal o por
instilacin nasal. Al segundo o tercer das posdesafo
se evala el grado de proteccin como el porcentaje de
aves que no se enferman segn un criterio establecido.
Un criterio estricto es considerar que un ave est
enferma cuando presenta signos clnicos de coriza
infecciosa, mucus en uno o ambos senos paranasales
y aislamiento de H. paragallinarum de uno o ambos
senos paranasales. 87,88 Adems, los signos de coriza
se pueden clasificar en cuatro grados; en un criterio
estricto slo se consideran con coriza (positivas) los
grados 2, 3 y 4 (Figuras 3 y 4).
Es ampliamente reconocido que los tres serogrupos
de Blackall representan tres inmunovariedades
diferentes. 89 En general, el dogma aceptado es que
no existe proteccin cruzada entre las serovariedades
de estos serogrupos. Recientemente, Soriano et al.45
evaluaron la proteccin cruzada en aves inmunizadas
y desafiadas con las cepas de referencia de H.
paragallinarum en el esquema de Blackall. Los
resultados obtenidos en este trabajo confirmaron
que las tres inmunovariedades son diferentes. Sin
embargo, se observ cierta proteccin cruzada entre las
serovariedades de los tres serogrupos. Estos resultados
permitirn guiar estrategias en la prevencin de la
coriza infecciosa, ya que en la actualidad se emplean
bacterinas comerciales bivalentes (serovariedades A-1
y C-1) y trivalentes (A-1, B-1 y C-2) contra esta
enfermedad en todo el mundo. En este sentido,
hasta la fecha existan discrepancias en cuanto a
la proteccin conferida por bacterinas bivalentes
contra aislamientos serovariedad B. 69,90 Sin embargo,
varios trabajos confirman la necesidad de usar
bacterinas trivalentes en todas las reas donde se han
diagnosticado cepas regionales de la serovariedad
B. Por ejemplo, en un estudio reciente en el cual
se emplearon cepas de referencia, se inform que
una bacterina bivalente no confiri proteccin en
aves desafiadas con un aislamiento serovariedad
B-1 de Mxico. 91 En Argentina, de manera similar
pero empleando cepas regionales, Terzolo et al.88
mostraron que bacterinas comerciales bivalentes no
protegieron a pollos desafiados con un aislamiento
de la serovariedad B. En otros ensayos, Yamaguchi
et al.92 informaron que las bacterinas bivalentes
proporcionaron proteccin contra la cepa Spross (B),

Vet. Mx., 35 (3) 2004

273

characterization of B isolates, as notorious antigenic

differences among B serovar strains exists.

Treatment and disinfection

Several chemotherapeutics and antibiotics have been
used for the treatment of infectious coryza:
streptomycin, 95 spectinomycin, streptomycin-spectinomycin 96 or sulphachloropiridazine-trimetroprim 97 and
sulphadimetoxin-trimetroprim, 98 among other combinations. Norfloxacin nicotinate 99 and enrofloxacin100
quinolones have showed excellent results in the
treatment of this disease. Broilers experimentally
infected with infectious coryza were treated with
enrofloxacin and bromhe-xine chlorhydrate or with
enrofloxacin alone. 80 A reduction in clinical signs
was observed in chickens that received the combined
treatment but not with enrofloxacin alone.
Furthermore, in comparison to an untreated control
group, a higher weight mean difference of 194 g was
obtained in chickens that have received the combined
treatment.
A reduction of the infectious coryza impact was
reported by Bragg,101 who used didecildimetylamonium chloride as part of disinfection programs in
both layer and broiler farms. Application of this
disinfectant procedure provoked less severe clinical
signs and shorter disease course in both vaccinated
and non-vaccinated chickens challenged with A-1,
B-1, C-2, and C-3 isolates of H. paragallinarum. Other
disinfectants for controlling infectious coryza have
not been studied yet.
Antimicrobial resistance of H. paragallinarum to
currently used antibiotics and chemotherapeutics may
be rapidly acquired and this phenomenon has to be
cautiously followed up. For this reason, sensibility tests
for the selection of the most suitable antimicrobial
agents, must included the regional isolates involved in
each outbreak.102 In addition it has to be remembered
that any treatment may partially control an infection
but is unable to completely eliminate it from a diseased
flock, therefore associated disinfection programs are
very important to be accomplished. As any bird which
recovers from the disease may act as a healthy carrier,
it is highly recommended, in every exposed farms, first
to treat the affected animals and afterwards vaccinate
all new incoming birds immediately after arrival into
the infected hen house.

Conclusion

pero fallaron contra otras dos cepas B sudafricanas.

De forma anloga, Terzolo et al.88 demostraron
que una bacterina trivalente elaborada con una
cepa B de referencia fall en pollos inmunizados
y posteriormente desafiados con una cepa regional
de la serovariedad B. Bowles et al.,93 en un estudio
comparativo muy amplio, determinaron la diversidad
gentica y las relaciones entre 118 cepas provenientes
de seis continentes, mediante movilidad electrofortica
de ocho enzimas metablicas codificadas por genes
cromosomales. Es interesante destacar que en este
trabajo, las cepas de H. paragallinarum estudiadas se
clasificaron en dos grupos heterogneos, mientras
que nueve cepas regionales de la serovariedad B
de Argentina, ocuparon un grupo diferente, lo que
demuestra que stas estn genticamente distantes
de las otras cepas estudiadas, independientemente de
la serovariedad a la cual pertenecen. Es decir que
las cepas B de Argentina fueron incluso diferentes
a otras cepas B de diversas regiones del mundo.
Recientemente Jacobs et al.94 informaron que pollos
inmunizados con una bacterina comercial trivalente
mostraron proteccin al desafo con un aislamiento
de Ecuador serovariedad C, pero no cuando fueron
desafiados con aislamientos B procedentes de varios
pases (Ecuador, Argentina, Estados Unidos de
Amrica y Zimbabwe). Los aislamientos fueron
reconocidos por antisueros elaborados para las
cepas Spross (B) y H-18 (C). La proteccin contra
estos aislamientos B se increment cuando un
aislamiento B de Ecuador fue incluido en la bacterina
trivalente. Con base en estos hallazgos, los autores
consideran que estos aislamientos B (variantes)
representan una nueva inmunovariedad, designando
al inmungeno que incluye este aislamiento como
bacterina tetravalente. El criterio utilizado por estos
autores en su investigacin no es suficiente para
proponer la existencia de una nueva inmunovariedad.
Es muy probable que estos aislamientos constituyan
una nueva, o nuevas, serovariedades dentro del
serogrupo B, de forma similar a los serogrupos A o
C que incluyen cuatro serovariedades cada uno. Los
resultados obtenidos sealan la necesidad de realizar
estudios de caracterizacin de hemoaglutininas, ya
que evidencian que existen notables diferencias
inmunolgicas entre diferentes cepas de la
serovariedad B. Es necesario incluir cepas regionales
de la serovariedad B en la formulacin de bacterinas
locales.

Tratamiento y desinfeccin
Infectious coryza outbreaks could be produced by
both NAD- dependent or independent isolates of
H. paragallinarum. High morbidity with increased
mortality of affected birds could be recorded.

274

En el tratamiento de la coriza infecciosa se han

empleado varios quimioteraputicos y antibiticos
como:
estreptomicina, 95
espectinomicina,
la

Vaccination of susceptible chicken flocks has to be

performed with trivalent vaccines including serovars
A-1, B-1 and C-2, being the main preventive strategy
against infectious coryza. New independent studies
advises the usage of tetravalent vaccines, particularly
in such geographical areas in which variant serovar
B outbreaks are prevalent. In some cases, local
isolates included into any these vaccines could
greatly increase protection. Serological classification
of H. paragallinarum isolates based on the Blackalls
scheme is of paramount importance. The results
summarized in this review could be used as a
guide to standardize, both epizootiological studies
and prophylactic practices against infectious coryza,
carried either in Mexico or elsewhere.

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combinacin estreptomicina-espectinomicina 96 o las

combinaciones sulfacloropiridazina-trimetroprim 97 y
sulfadimetoxina-trimetroprim, 98 entre otras. Las
quinolonas
nicotinato
de
norfloxacina 99
y
100
enrofloxacina han dado excelentes resultados en
el tratamiento de esta enfermedad. Se inform
del tratamiento de pollos de engorda con coriza
infecciosa, de manera experimental, empleando
enrofloxacina y clorhidrato de bromhexina. 80 Los
resultados obtenidos mostraron que los pollos que
recibieron la combinacin redujeron el tiempo de
signos clnicos, a diferencia de los que recibieron
nicamente enrofloxacina. Adems, la diferencia en
peso del grupo de aves que recibi la combinacin
estuvo 194 g por arriba del grupo testigo.
Bragg101 inform del uso de cloruro de
didecildimetilamonio en programas de desinfeccin
continuos en granjas de gallinas de postura y pollos
de engorda, reduciendo el impacto de la coriza
infecciosa. Menciona que pollos inmunizados y no
inmunizados contra la coriza infecciosa, y desafiados
con H. paragallinarum de las serovariedades A-1, B-1,
C-2 y C-3, mostraron signos menos severos y un curso
ms corto de la enfermedad. Se desconoce el efecto
de otros desinfectantes en el control de la coriza
infecciosa.
Se debe considerar que H. paragallinarum puede
generar resistencia a los antibiticos y quimioterapeticos empleados actualmente. Por ello es necesario
realizar pruebas de sensibilidad para seleccionar el
antimicrobiano ms adecuado para tratar a la cepa
actuante en un determinado brote.102 Tambin debe
considerarse que luego del tratamiento, en las granjas
afectadas por la enfermedad, la infeccin puede
controlarse pero nunca se elimina totalmente, siendo
importantes los programas de desinfeccin. Como las
aves actan como portadoras, en casos de brotes en
granjas con edades mltiples, lo ms recomendable
es tratar en primera instancia y adems indicar la
inmunizacin de todas las nuevas aves que ingresen al
establecimiento afectado.

Conclusiones
Brotes de coriza infecciosa pueden ser ocasionados
por H. paragallinarum tanto dependientes como
independientes de NAD. Se puede registrar morbilidad
alta con incremento en la mortalidad de aves afectadas.
La produccin de huevo en gallinas de postura es
afectada considerablemente. La bacterinizacin de
parvadas susceptibles con inmungenos trivalentes
que incluyan las serovariedades A-1, B-1 y C-2 es la
estrategia principal en la prevencin de la coriza
infecciosa. La inclusin de aislamientos locales en
estos productos puede incrementar la proteccin,

Vet. Mx., 35 (3) 2004

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